Platelet Factor 4 Activity and Thromboembolic Episodes TAKASHI OKUNO, M.D., AND DALE CROCKATT, B.S.
THERE HAVE BEEN VARIOUS laboratory procedures for the diagnosis of intravascular coagulopathy and hypercoagulability. Determination of antithrombin III activity is one such procedure. Its correlations with clinical values have been extensively studied and appear to be well established in clinical laboratories. 410 Another procedure for the possible detection of thrombotic episodes, determination of platelet factor 4 activity, was recently described. 7,9,11 A given level of platelet factor 4 was found to be closely related to platelet destruction or platelet consumption, and its activity seems to be related to thromboembolic episodes. According to these reports, many patients who had recently had thrombosis, including disseminated intravascular coagulopathy and myocardial infarction, had elevated platelet factor 4 activities. We have studied the value of determination of platelet factor 4 activity in management of patients with various diseases. Materials and Methods We studied a total of 323 patients hospitalized at Lutheran General Hospital from June 1975 to August Received February 12, 1976; received revised manuscript May 24, 1976; accepted for publication June 10, 1976. Address reprint requests to Dr. Okuno: Lutheran General Hospital, 1775 Dempster Street, Park Ridge, Illinois 60068.
Section of Hematology, Lutheran General Hospital, Park Ridge, Illinois, and University of Illinois, Chicago, Illinois
1975. Additionally, 43 donors to the blood bank were studied as controls. The patients were divided into groups according to their diseases. Fifty-five patients were diagnosed as alcoholics, but had no history of thromboemboli, myocardial infarction, low platelet counts, hepatic disease, or receiving estrogen in the preceding four months; 52 patients had platelet counts below 100,000/cu mm; 43 patients had the diagnosis of thromboembolism or histories of thromboembolism within four months of the study, but no diagnosis of recent myocardial infarction, hepatic disease, diabetes mellitus, or alcoholism; 39 patients were diagnosed as having diabetes mellitus; 24 patients had arteriosclerotic heart disease or angina pectoris, but no evidence of myocardial infarction; 20 patients had hepatic diseases; 13 patients were receiving estrogen therapy; ten patients were taking oral contraceptives; eight patients had neoplasms but had normal platelet counts; 36 patients had other diseases. For quantitative assay of platelet factor 4, a heparin thrombin clotting time method, that of Fuster and associates, 7 was used. In brief, a test plasma was treated by centrifugation and heating to render it platelet-poor and free of fibrinogen, heparin cofactor and other proteins. To the treated plasma, heparin, platelet-poor plasma, and thrombin were added and the Fibrometer (BBL, Baltimore, Md.) clotting time was recorded. For quantitative assay of antithrombin III, the method of Biggs and co-workers was used. 4 In brief, the heated test plasma was incubated with thrombin. After incubation, the mixture was transferred to a fibrinogen solution and the clotting times were recorded. Platelet counts and other hematologic studies were done by conventional methods.
351
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Okuno, Takashi, and Crockatt, Dale: Platelet factor 4 activity and thromboembolic episodes. Am J Clin Pathol 67: 351-355, 1977. Platelet factor 4 activity was evaluated in 323 patients who had various diseases. High incidences (30% or more) of elevated levels of platelet factor 4 were found in patients with recent myocardial infarctions, recent episodes of thromboembolism, hepatic diseases, and low platelet counts. Three of the four patients who had prosthetic heart valves had markedly elevated platelet factor 4 activities. It appears that elevation of platelet factor 4 activity is associated with a high platelet turnover such as is seen in intravascular coagulation or thromboembolism. Determination of platelet factor 4 activity may be a valuable laboratory tool for the diagnosis of thromboembolism. (Key words: Antithrombin HI activity; Intravascular coagulation; Liver disease; Platelet factor 4; Thromboembolism.)
OKUNO AND CROCKATT
352
Discussion
Table 1. Age and Sex Distribution of Patients Studied
Recent myocardial infarction Low platelet count Thrombosis Hepatic disease Diabetes With thrombosis Without thrombosis Alcoholism Contraceptives Estrogen therapy Control
Male
Female
Total
58.2 (13)* 52.3 (28) 57.7 (18) 45.4(11) 58.1 (23) 59.2 (7) 57.6 (16) 37.7 (38)
66.8 (11) 52.4 (24) 58.0 (25) 59.7 (9) 63.8(16) 61.5 (9) 66.7 (7) 45.7(17) 29.0(10) 54.5(13) 42.4(14)
62.0 (24) 52.3 (52) 57.9 (43) 51.8(20) 60.4 (39) 60.4 (16) 60.4 (23) 40.2 (55) 29.0 (10) 54.5 (13) 38.7 (43)
— — 37.3 (29)
* Number of patients.
Platelet factor 4, a heparin-neutralizing substance, is a heat-stable protein with a low molecular weight. It is released from platelets during the initial stages of blood coagulation.5,7,s Because of its heat-resistant characteristics, platelet factor 4 can be differentiated from other antithrombin activities in plasma by heating test plasma to 60-65 C for 10 minutes.8 The significance of platelet factor 4 is somewhat unclear. Recent reports, however, indicate that increased platelet factor 4 activity is closely related to thromboembolic phenomena.2'9,11'12 Since platelet factor 4 is liberated by blood coagulation and its activity appears to reflect the rate of platelet consumption, it has been suggested that chronic as well as acute episodes of intravascular coagulation can be detected by measuring its activity in plasma. Fuster and co-workers also demonstrated in dogs that experimentally induced intravascular coagulation drastically increased platelet factor 4 activity.8 Table 2. Normal Platelet Factor 4 and Antithrombin III Values
Mean Standard deviation Normal range
Platelet Factor 4 (n = 43) (%)
Antithrombin III
100.4 9.0 82.4-118.4
99.9 3.7 92.5-107.3
(n = 43) (%)
Table 3. Platelet Factor 4 and Antithrombin III Values in the Diseases Studied
Recent myocardial infarction Low platelet count Thrombosis Hepatic disease Diabetes With thrombosis Without thrombosis Alcoholism Estrogen therapy Arteriosclerotic heart disease Neoplasms with normal platelet count Contraceptives Control
Platelet Factor 4 Positive
Antithrombin III Positive
(%)
(%)
No. Patients Studied
33.3 32.7 30.2 30.0 17.9 25.0 13.0 14.5 7.7
41.7 32.7 67.4 50.0 51.3 68.8 39.1 3.6 23.0
24 52 43 20 39 16 23 55 13
0.4
8.7
23
12.5 0 2.3
12.5 90.0 0
8 10 43
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Results The age and sex distribution of the patients is shown in Table 1. On the average, controls were younger than patient groups, but the sexes were equally represented in all groups. The means and normal ranges of platelet factor 4 and antithrombin III, based on values from 43 blood donors and healthy hospital employees, are shown in Table 2. Results more than two standard deviations (SD) from normal values were considered abnormal. There was no significant difference between values obtained from male and female control subjects. Only one blood donor had elevated platelet factor 4 activity, while all antithrombin III results were within the normal range. The platelet factor 4 and antithrombin III values found in various groups of patients are shown in Table 3. High incidences of elevated platelet factor 4 activity were found in various thrombotic diseases. None of the patients taking oral contraceptives had elevated platelet factor 4 activity, but 90% of them had increased antithrombin III levels (Fig. 1). Platelet factor 4 and antithrombin III activities in the thrombotic group were further analyzed according to the sites of thromboembolism (Table 4). The sites of thrombosis did not appear to relate significantly to the values of platelet factor 4 or antithrombin III. The results for patients who had low platelet counts are shown in Table 5. Many patients who had cirrhosis of the liver and neoplasms were shown to have increased platelet factor 4 activities. Both of the patients who had disseminated intravascular coagulation had elevated platelet factor 4 levels, while neither of the patients who had idiopathic thrombocytopenic purpura had increased platelet factor 4. Four patients had prosthetic heart valves. Their results are shown in Table 6. One of the patients, a 35-year-old woman, had episodes of multiple thromboemboli in abdominal organs.
A.J.C.P. . April 1977
Vol. 67 • No. 4
353
PLATELET FACTOR 4 ACTIVITY
Antithrombin activities, in contrast to platelet factor 4, have been extensively studied. The procedure most often used in clinical laboratories for their determination is measurement of thrombin activity after incubation of a mixture of thrombin and test serum.4 The close correlation of antithrombin activity with thrombotic episodes appears to be established, and many diseases associated with thromboembolism are identified by low antithrombin activities.l,4,1° Our studies of 323 hospitalized patients appear to support the value of platelet factor 4 determination in management of patients suspected to have thromboembolism. About 30% of the patients in a thromboembolic group had increased platelet factor 4 activity (Table 3). This group consisted of patients who had had thrombotic episodes within four months of the study. There was no significant difference
100 90 60 - >0
-••
so - 50 - -
30 - 20 -10 --
0
>§
C Site Leg Lung Brain Multiple TOTAL
Platelet Factor 4
Antithrombin III
Total
6(31.6%) 4 (25.0%) 2 (28.0%) 1
15 (78.9%) 10 (62.5%) 3 (42.9%) 1
19 16 7 1
13 (30.2%)
29 (67.4%)
43
T3
Table 5. Relation between Thrombocytopenia with Various Diseases and Platelet Factor 4 and Antithrombin III Activities Positive by
Cirrhosis of the liver Neoplasms Diabetes Myocardial infarction Alcoholism Fractures Idiopathic thrombocytopenic purpura Disseminated intravascular coagulopathy Gastrointestinal bleeding Thrombosis Prosthetic heart valves Others TOTAL
Platelet Factor 4
Antithrombin III
4 (28.6%) 4 (33.3%) 1 3 (75.0%) 1 0
8(57.1%) 2 (16.7%) 1 3 (75.0%) 1 0
0
1
2
2 0 1 1 0
0 0 1 0 0
2 2 2 1 2
17 (32.7%)
17 (32.7%)
No. Patients Studied 14 12 5 4 3 3
52
_1
Q. E.
9
1
FIG. 1. Positive results in platelet factor 4 (Pf-4) and antithrombin III (AT-III) activities among patients with various diseases.
relating to the sites of thromboembolism (Table 4). Many patients who had recently had myocardial infarctions also had elevated platelet factor 4 activity. The high incidence in this group is in sharp contrast to that in the arteriosclerotic heart disease group, in which the incidence of abnormal results was only 0.4% among 23 patients studied. The difference was statistically significant (x 2 = 6.373, p < 0.02). Age and sex distributions were found to be comparable in the two groups (Table 1). The close relationship between diabetes mellitus and thromboembolism has been known for a long time. Antithrombin III activity has been reported to be decreased in maturity-onset diabetes.3 Platelet factor 4 activity was recently found to be increased in diabetics, and it was thought that increased platelet factor 4 activity might play an important role in pathogenesis of thromboembolic complications.5 In our studies, about 18% of the diabetic patients had abnormally increased platelet factor 4 activity. The increase was
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 5, 2016
Table 4. Relation between sites of Thromboembolism and Platelet Factor 4 and Antithrombin III Activities
%
OKUNO AND CROCKATT
354
Table 6. Platelet Factor 4 Deviations from Normal and Various Diseases Platelet Factor 4 Deviation No Patients Studied
Total Abnormal
24
8
5
2
1
52 43 20
16 13 4
1 3 0
8 7 1
7 3 4
39
6
0
3
3
Recent myocardial infarction Low platelet count Thrombosis Hepatic disease Diabetes mellitus
ModSlight* erate*
Marked*
• Slight. 118.5-127.4%: moderate. 127.5-136.4%: marked, more than 136.5%.
Patient's Age (yr.), Sex 55, 51, 59, 35,
M M F Ft
Platelet Factor 4
Antithrombin III
(%) 146* 125* 93 150*
(%)
Platelet Count (/cu mm)
Type of Prosthesis
103 99 107 95
308,000 115,000 343,000 290,000
Aortic Aortic Mitral Mitral
* Abnormal results. t Had had many episodes of thromboembolism in abdominal organs.
particularly great in patients who had had thrombotic episodes, although there was no statistically significant difference between patients who had had thrombotic episodes and those without thrombotic episodes (X2 = 0.92, p > 0.50). Since platelet factor 4 activity appears to be related to platelet turnover, we also studied patients who had low platelet counts, classified according to their diseases (Table 5). A substantial number of these patients with cirrhosis of the liver had increased platelet factor 4 activity. Both patients who had disseminated intravascular coagulopathy had increased platelet factor 4 activity, while their antithrombin III activity was normal. Neither of two patients who had idiopathic thrombocytopenic purpura had increased platelet factor 4 activity. Since most patients who have idiopathic thrombocytopenic purpura have reduced platelet survival and high platelet turnover rates, reduced platelet factor 4 activity in such patients is surprising. Similar observations of reduced activity were made by O'Brien and associates.11 Thus, none of the eight patients who had idiopathic thrombo-
cytopenic purpura had increased platelet factor 4 activity, but rather, their platelet factor 4 activity was markedly reduced. As a possible explanation for this, they postulated that immunologic platelet destruction occurring in the spleen and liver, as in idiopathic thrombocytopenic purpura, may not liberate platelet factor 4 into the circulating blood. The release of platelet factor 4 in blood may occur only during intravascular platelet destruction, as seen in intravascular coagulopathy. The relatively high incidence of elevated platelet factor 4 activity in alcoholic patients in our studies is rather surprising. It may reflect reduced platelet survival, possibly due to folate deficiency, since it is known that alcohol interferes with platelet kinetics at various stages.6 Three of the 55 patients with alcoholism had thrombocytopenia, but low platelet counts did not appear to be related to high platelet factor 4 activity. An attempt was made to classify abnormal platelet factor 4 activities according to their deviation from the normal values. Thus, results that deviated by 2 or 3 standard deviations'(118.5-127.4%) are considered "slight" abnormal results; between 3 and 4 standard deviations (127.5-136.4%), "moderate" abnormal results; and more than 4 standard deviations (more than 136.5%), "marked" abnormal results. Table 6 shows the classification of platelet factor 4 results and various diseases. Although the myocardial infarction group appears to fall more into the "slight" abnormal results, most of the patients showed "moderate" to "marked" abnormalities. A high incidence of thromboembolic phenomena has been observed in patients who have prosthetic heart valves. Thrombus formation on a prosthetic valve is thought to be an origin of thromboembolic complications. Reduced platelet survival in such patients has also been demonstrated. Recent studies indicate that the shortened survival is due to direct damage by the prosthesis, as has been seen in hemolysis of erythrocytes.13 Four patients in our studies had prosthetic heart valves. Three of them had elevated platelet factor 4 activity, while antithrombin III levels were normal. All three patients had moderately to markedly abnormal platelet factor 4 activities. One of the patients, though not thrombocytopenic, had had numerous episodes of thromboembolism in abdominal organs (Table 7). References 1. Abildgaard U: Heparin cofactor and antithrombin III. Thromb Diath Haemorrh 33:38-42, 1974 2. Aloborujeh M, Unterharnscheidt W, Riegel K: The activity of
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Table 7. Prosthetic Heart Valves and Platelet Factor 4 and Antithrombin III Activities
A.J.C.P. • April 1977
Vol. 67 • No. 4
3. 4. 5. 6. 7.
PLATELET FACTOR 4 ACTIVITY
platelet factor IV in plasma of healthy and high risk newborn. Z Kinderheilk 116:137-142, 1974 Banerjee RN, Jahni AL, Kumar V, et al: Antithrombin III deficiency in maturity onset diabetes mellitus and arteriosclerosis. Thromb Diath Haemorrh 31:339-345, 1974 Biggs R, Deuson W, Akman N, et al: Antithrombin III, Antifactor Xa and Heparin. Br J Hematol 19:283-305, 1970 Chmielewski J, Farbiszewski R: Platelet factor 4 (Pf-4) release during human platelet aggregation in diabetic patients. Thromb Diath Haemorrh 24:203-205, 1970 Eichner ER: The hematologic disorders of alcoholism. Am J Med 54:621-630, 1973 Fuster V, Bowie EW, Kazmier FJ, et al: Platelet consumption in chronically induced plasma platelet factor 4-like activity reflecting rate of intravascular coagulation in dogs. Thromb Res 4:247-260, 1974
355
8. Fuster V, Kazmier F, Bowie EJW, et al: Assay of platelet factor 4 in plasma. Mayo Clin Proc 48:103-106. 1973 9. Green PJ: The heparin thrombin clotting time. Lancet 1: 799, 1975 10. Hender U, Nilsson IM: Antithrombin-III in a clinical material. Bibl Anat 12:267-271, 1973 11. O'Brien JR, Etherington M, Jamieson S, et al: Heparin thrombin clotting time and platelet factor 4. Lancet ii: 656-657, 1974 12. O'Brien JR, Tulenski VG, Etherington M, et al: Platelet function studies before and after operation and the effect of postoperative thrombosis. J Lab Clin Med 83:342, 1974 13. Stuart RK, McDonald JW, Ahuja SP. et al: Platelet survival in patients with prosthetic heart valves. Am J Cardiol 33:840-844, 1974
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THERE HAVE BEEN VARIOUS laboratory procedures for the diagnosis of intravascular coagulopathy and hypercoagulability. Determination of antithrombin III activity is one such procedure. Its correlations with clinical values have been extensively studied and appear to be well established in clinical laboratories. 410 Another procedure for the possible detection of thrombotic episodes, determination of platelet factor 4 activity, was recently described. 7,9,11 A given level of platelet factor 4 was found to be closely related to platelet destruction or platelet consumption, and its activity seems to be related to thromboembolic episodes. According to these reports, many patients who had recently had thrombosis, including disseminated intravascular coagulopathy and myocardial infarction, had elevated platelet factor 4 activities. We have studied the value of determination of platelet factor 4 activity in management of patients with various diseases. Materials and Methods We studied a total of 323 patients hospitalized at Lutheran General Hospital from June 1975 to August Received February 12, 1976; received revised manuscript May 24, 1976; accepted for publication June 10, 1976. Address reprint requests to Dr. Okuno: Lutheran General Hospital, 1775 Dempster Street, Park Ridge, Illinois 60068.
Section of Hematology, Lutheran General Hospital, Park Ridge, Illinois, and University of Illinois, Chicago, Illinois
1975. Additionally, 43 donors to the blood bank were studied as controls. The patients were divided into groups according to their diseases. Fifty-five patients were diagnosed as alcoholics, but had no history of thromboemboli, myocardial infarction, low platelet counts, hepatic disease, or receiving estrogen in the preceding four months; 52 patients had platelet counts below 100,000/cu mm; 43 patients had the diagnosis of thromboembolism or histories of thromboembolism within four months of the study, but no diagnosis of recent myocardial infarction, hepatic disease, diabetes mellitus, or alcoholism; 39 patients were diagnosed as having diabetes mellitus; 24 patients had arteriosclerotic heart disease or angina pectoris, but no evidence of myocardial infarction; 20 patients had hepatic diseases; 13 patients were receiving estrogen therapy; ten patients were taking oral contraceptives; eight patients had neoplasms but had normal platelet counts; 36 patients had other diseases. For quantitative assay of platelet factor 4, a heparin thrombin clotting time method, that of Fuster and associates, 7 was used. In brief, a test plasma was treated by centrifugation and heating to render it platelet-poor and free of fibrinogen, heparin cofactor and other proteins. To the treated plasma, heparin, platelet-poor plasma, and thrombin were added and the Fibrometer (BBL, Baltimore, Md.) clotting time was recorded. For quantitative assay of antithrombin III, the method of Biggs and co-workers was used. 4 In brief, the heated test plasma was incubated with thrombin. After incubation, the mixture was transferred to a fibrinogen solution and the clotting times were recorded. Platelet counts and other hematologic studies were done by conventional methods.
351
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Okuno, Takashi, and Crockatt, Dale: Platelet factor 4 activity and thromboembolic episodes. Am J Clin Pathol 67: 351-355, 1977. Platelet factor 4 activity was evaluated in 323 patients who had various diseases. High incidences (30% or more) of elevated levels of platelet factor 4 were found in patients with recent myocardial infarctions, recent episodes of thromboembolism, hepatic diseases, and low platelet counts. Three of the four patients who had prosthetic heart valves had markedly elevated platelet factor 4 activities. It appears that elevation of platelet factor 4 activity is associated with a high platelet turnover such as is seen in intravascular coagulation or thromboembolism. Determination of platelet factor 4 activity may be a valuable laboratory tool for the diagnosis of thromboembolism. (Key words: Antithrombin HI activity; Intravascular coagulation; Liver disease; Platelet factor 4; Thromboembolism.)
OKUNO AND CROCKATT
352
Discussion
Table 1. Age and Sex Distribution of Patients Studied
Recent myocardial infarction Low platelet count Thrombosis Hepatic disease Diabetes With thrombosis Without thrombosis Alcoholism Contraceptives Estrogen therapy Control
Male
Female
Total
58.2 (13)* 52.3 (28) 57.7 (18) 45.4(11) 58.1 (23) 59.2 (7) 57.6 (16) 37.7 (38)
66.8 (11) 52.4 (24) 58.0 (25) 59.7 (9) 63.8(16) 61.5 (9) 66.7 (7) 45.7(17) 29.0(10) 54.5(13) 42.4(14)
62.0 (24) 52.3 (52) 57.9 (43) 51.8(20) 60.4 (39) 60.4 (16) 60.4 (23) 40.2 (55) 29.0 (10) 54.5 (13) 38.7 (43)
— — 37.3 (29)
* Number of patients.
Platelet factor 4, a heparin-neutralizing substance, is a heat-stable protein with a low molecular weight. It is released from platelets during the initial stages of blood coagulation.5,7,s Because of its heat-resistant characteristics, platelet factor 4 can be differentiated from other antithrombin activities in plasma by heating test plasma to 60-65 C for 10 minutes.8 The significance of platelet factor 4 is somewhat unclear. Recent reports, however, indicate that increased platelet factor 4 activity is closely related to thromboembolic phenomena.2'9,11'12 Since platelet factor 4 is liberated by blood coagulation and its activity appears to reflect the rate of platelet consumption, it has been suggested that chronic as well as acute episodes of intravascular coagulation can be detected by measuring its activity in plasma. Fuster and co-workers also demonstrated in dogs that experimentally induced intravascular coagulation drastically increased platelet factor 4 activity.8 Table 2. Normal Platelet Factor 4 and Antithrombin III Values
Mean Standard deviation Normal range
Platelet Factor 4 (n = 43) (%)
Antithrombin III
100.4 9.0 82.4-118.4
99.9 3.7 92.5-107.3
(n = 43) (%)
Table 3. Platelet Factor 4 and Antithrombin III Values in the Diseases Studied
Recent myocardial infarction Low platelet count Thrombosis Hepatic disease Diabetes With thrombosis Without thrombosis Alcoholism Estrogen therapy Arteriosclerotic heart disease Neoplasms with normal platelet count Contraceptives Control
Platelet Factor 4 Positive
Antithrombin III Positive
(%)
(%)
No. Patients Studied
33.3 32.7 30.2 30.0 17.9 25.0 13.0 14.5 7.7
41.7 32.7 67.4 50.0 51.3 68.8 39.1 3.6 23.0
24 52 43 20 39 16 23 55 13
0.4
8.7
23
12.5 0 2.3
12.5 90.0 0
8 10 43
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 5, 2016
Results The age and sex distribution of the patients is shown in Table 1. On the average, controls were younger than patient groups, but the sexes were equally represented in all groups. The means and normal ranges of platelet factor 4 and antithrombin III, based on values from 43 blood donors and healthy hospital employees, are shown in Table 2. Results more than two standard deviations (SD) from normal values were considered abnormal. There was no significant difference between values obtained from male and female control subjects. Only one blood donor had elevated platelet factor 4 activity, while all antithrombin III results were within the normal range. The platelet factor 4 and antithrombin III values found in various groups of patients are shown in Table 3. High incidences of elevated platelet factor 4 activity were found in various thrombotic diseases. None of the patients taking oral contraceptives had elevated platelet factor 4 activity, but 90% of them had increased antithrombin III levels (Fig. 1). Platelet factor 4 and antithrombin III activities in the thrombotic group were further analyzed according to the sites of thromboembolism (Table 4). The sites of thrombosis did not appear to relate significantly to the values of platelet factor 4 or antithrombin III. The results for patients who had low platelet counts are shown in Table 5. Many patients who had cirrhosis of the liver and neoplasms were shown to have increased platelet factor 4 activities. Both of the patients who had disseminated intravascular coagulation had elevated platelet factor 4 levels, while neither of the patients who had idiopathic thrombocytopenic purpura had increased platelet factor 4. Four patients had prosthetic heart valves. Their results are shown in Table 6. One of the patients, a 35-year-old woman, had episodes of multiple thromboemboli in abdominal organs.
A.J.C.P. . April 1977
Vol. 67 • No. 4
353
PLATELET FACTOR 4 ACTIVITY
Antithrombin activities, in contrast to platelet factor 4, have been extensively studied. The procedure most often used in clinical laboratories for their determination is measurement of thrombin activity after incubation of a mixture of thrombin and test serum.4 The close correlation of antithrombin activity with thrombotic episodes appears to be established, and many diseases associated with thromboembolism are identified by low antithrombin activities.l,4,1° Our studies of 323 hospitalized patients appear to support the value of platelet factor 4 determination in management of patients suspected to have thromboembolism. About 30% of the patients in a thromboembolic group had increased platelet factor 4 activity (Table 3). This group consisted of patients who had had thrombotic episodes within four months of the study. There was no significant difference
100 90 60 - >0
-••
so - 50 - -
30 - 20 -10 --
0
>§
C Site Leg Lung Brain Multiple TOTAL
Platelet Factor 4
Antithrombin III
Total
6(31.6%) 4 (25.0%) 2 (28.0%) 1
15 (78.9%) 10 (62.5%) 3 (42.9%) 1
19 16 7 1
13 (30.2%)
29 (67.4%)
43
T3
Table 5. Relation between Thrombocytopenia with Various Diseases and Platelet Factor 4 and Antithrombin III Activities Positive by
Cirrhosis of the liver Neoplasms Diabetes Myocardial infarction Alcoholism Fractures Idiopathic thrombocytopenic purpura Disseminated intravascular coagulopathy Gastrointestinal bleeding Thrombosis Prosthetic heart valves Others TOTAL
Platelet Factor 4
Antithrombin III
4 (28.6%) 4 (33.3%) 1 3 (75.0%) 1 0
8(57.1%) 2 (16.7%) 1 3 (75.0%) 1 0
0
1
2
2 0 1 1 0
0 0 1 0 0
2 2 2 1 2
17 (32.7%)
17 (32.7%)
No. Patients Studied 14 12 5 4 3 3
52
_1
Q. E.
9
1
FIG. 1. Positive results in platelet factor 4 (Pf-4) and antithrombin III (AT-III) activities among patients with various diseases.
relating to the sites of thromboembolism (Table 4). Many patients who had recently had myocardial infarctions also had elevated platelet factor 4 activity. The high incidence in this group is in sharp contrast to that in the arteriosclerotic heart disease group, in which the incidence of abnormal results was only 0.4% among 23 patients studied. The difference was statistically significant (x 2 = 6.373, p < 0.02). Age and sex distributions were found to be comparable in the two groups (Table 1). The close relationship between diabetes mellitus and thromboembolism has been known for a long time. Antithrombin III activity has been reported to be decreased in maturity-onset diabetes.3 Platelet factor 4 activity was recently found to be increased in diabetics, and it was thought that increased platelet factor 4 activity might play an important role in pathogenesis of thromboembolic complications.5 In our studies, about 18% of the diabetic patients had abnormally increased platelet factor 4 activity. The increase was
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 5, 2016
Table 4. Relation between sites of Thromboembolism and Platelet Factor 4 and Antithrombin III Activities
%
OKUNO AND CROCKATT
354
Table 6. Platelet Factor 4 Deviations from Normal and Various Diseases Platelet Factor 4 Deviation No Patients Studied
Total Abnormal
24
8
5
2
1
52 43 20
16 13 4
1 3 0
8 7 1
7 3 4
39
6
0
3
3
Recent myocardial infarction Low platelet count Thrombosis Hepatic disease Diabetes mellitus
ModSlight* erate*
Marked*
• Slight. 118.5-127.4%: moderate. 127.5-136.4%: marked, more than 136.5%.
Patient's Age (yr.), Sex 55, 51, 59, 35,
M M F Ft
Platelet Factor 4
Antithrombin III
(%) 146* 125* 93 150*
(%)
Platelet Count (/cu mm)
Type of Prosthesis
103 99 107 95
308,000 115,000 343,000 290,000
Aortic Aortic Mitral Mitral
* Abnormal results. t Had had many episodes of thromboembolism in abdominal organs.
particularly great in patients who had had thrombotic episodes, although there was no statistically significant difference between patients who had had thrombotic episodes and those without thrombotic episodes (X2 = 0.92, p > 0.50). Since platelet factor 4 activity appears to be related to platelet turnover, we also studied patients who had low platelet counts, classified according to their diseases (Table 5). A substantial number of these patients with cirrhosis of the liver had increased platelet factor 4 activity. Both patients who had disseminated intravascular coagulopathy had increased platelet factor 4 activity, while their antithrombin III activity was normal. Neither of two patients who had idiopathic thrombocytopenic purpura had increased platelet factor 4 activity. Since most patients who have idiopathic thrombocytopenic purpura have reduced platelet survival and high platelet turnover rates, reduced platelet factor 4 activity in such patients is surprising. Similar observations of reduced activity were made by O'Brien and associates.11 Thus, none of the eight patients who had idiopathic thrombo-
cytopenic purpura had increased platelet factor 4 activity, but rather, their platelet factor 4 activity was markedly reduced. As a possible explanation for this, they postulated that immunologic platelet destruction occurring in the spleen and liver, as in idiopathic thrombocytopenic purpura, may not liberate platelet factor 4 into the circulating blood. The release of platelet factor 4 in blood may occur only during intravascular platelet destruction, as seen in intravascular coagulopathy. The relatively high incidence of elevated platelet factor 4 activity in alcoholic patients in our studies is rather surprising. It may reflect reduced platelet survival, possibly due to folate deficiency, since it is known that alcohol interferes with platelet kinetics at various stages.6 Three of the 55 patients with alcoholism had thrombocytopenia, but low platelet counts did not appear to be related to high platelet factor 4 activity. An attempt was made to classify abnormal platelet factor 4 activities according to their deviation from the normal values. Thus, results that deviated by 2 or 3 standard deviations'(118.5-127.4%) are considered "slight" abnormal results; between 3 and 4 standard deviations (127.5-136.4%), "moderate" abnormal results; and more than 4 standard deviations (more than 136.5%), "marked" abnormal results. Table 6 shows the classification of platelet factor 4 results and various diseases. Although the myocardial infarction group appears to fall more into the "slight" abnormal results, most of the patients showed "moderate" to "marked" abnormalities. A high incidence of thromboembolic phenomena has been observed in patients who have prosthetic heart valves. Thrombus formation on a prosthetic valve is thought to be an origin of thromboembolic complications. Reduced platelet survival in such patients has also been demonstrated. Recent studies indicate that the shortened survival is due to direct damage by the prosthesis, as has been seen in hemolysis of erythrocytes.13 Four patients in our studies had prosthetic heart valves. Three of them had elevated platelet factor 4 activity, while antithrombin III levels were normal. All three patients had moderately to markedly abnormal platelet factor 4 activities. One of the patients, though not thrombocytopenic, had had numerous episodes of thromboembolism in abdominal organs (Table 7). References 1. Abildgaard U: Heparin cofactor and antithrombin III. Thromb Diath Haemorrh 33:38-42, 1974 2. Aloborujeh M, Unterharnscheidt W, Riegel K: The activity of
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Table 7. Prosthetic Heart Valves and Platelet Factor 4 and Antithrombin III Activities
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3. 4. 5. 6. 7.
PLATELET FACTOR 4 ACTIVITY
platelet factor IV in plasma of healthy and high risk newborn. Z Kinderheilk 116:137-142, 1974 Banerjee RN, Jahni AL, Kumar V, et al: Antithrombin III deficiency in maturity onset diabetes mellitus and arteriosclerosis. Thromb Diath Haemorrh 31:339-345, 1974 Biggs R, Deuson W, Akman N, et al: Antithrombin III, Antifactor Xa and Heparin. Br J Hematol 19:283-305, 1970 Chmielewski J, Farbiszewski R: Platelet factor 4 (Pf-4) release during human platelet aggregation in diabetic patients. Thromb Diath Haemorrh 24:203-205, 1970 Eichner ER: The hematologic disorders of alcoholism. Am J Med 54:621-630, 1973 Fuster V, Bowie EW, Kazmier FJ, et al: Platelet consumption in chronically induced plasma platelet factor 4-like activity reflecting rate of intravascular coagulation in dogs. Thromb Res 4:247-260, 1974
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8. Fuster V, Kazmier F, Bowie EJW, et al: Assay of platelet factor 4 in plasma. Mayo Clin Proc 48:103-106. 1973 9. Green PJ: The heparin thrombin clotting time. Lancet 1: 799, 1975 10. Hender U, Nilsson IM: Antithrombin-III in a clinical material. Bibl Anat 12:267-271, 1973 11. O'Brien JR, Etherington M, Jamieson S, et al: Heparin thrombin clotting time and platelet factor 4. Lancet ii: 656-657, 1974 12. O'Brien JR, Tulenski VG, Etherington M, et al: Platelet function studies before and after operation and the effect of postoperative thrombosis. J Lab Clin Med 83:342, 1974 13. Stuart RK, McDonald JW, Ahuja SP. et al: Platelet survival in patients with prosthetic heart valves. Am J Cardiol 33:840-844, 1974
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